Cross-linked gelatin composition comprising a wetting agent

ABSTRACT

Disclosed is a biocompatible, hemostatic, cross-linked gelatin composition comprising a biocompatible material comprising cross-linked gelatin and a sufficient amount of wetting agent to permit uniform wetting of the sponge in the presence of an aqueous solution.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No.13/477,541, filed on May 22, 2012, which is a continuation of U.S.application Ser. No. 10/068,812, filed on Feb. 4, 2002, now U.S. Pat.No. 8,187,625. This application claims priority to U.S. ProvisionalPatent Application Ser. No. 60/275,420, filed on Mar. 12, 2001, whichapplication is incorporated by reference herein in its entirety.

BACKGROUND OF INVENTION

1. Field of the Invention

This invention is directed to a biocompatible, hemostatic cross-linkedgelatin composition comprising cross-linked gelatin and a sufficientamount of wetting agent to permit uniform wetting of the gelatin in thepresence of an aqueous solution. This invention is further directedtowards methods of decreasing the hydration time of cross-linked gelatinby incorporating a biocompatible wetting agent into the gelatin. Thisinvention further relates to a kit of parts for preparing abiocompatible, hemostatic cross-linked gelatin composition comprising asterile syringe and non-hydrated, biocompatible, pledget comprising across-linked gelatin and a wetting agent.

2. References

The following patent applications and patents are cited and/orreferenced in this application as superscript numbers:

-   -   ¹Correll, et al., Proc. Soc. Exp. Biol. N.Y., 58:233 (1945).    -   ²Correll, et al., Surg. Gyn. and Obst., 82:585 (1945).    -   ³Correll, et al., U.S. Pat. No. 2,465,357, Therapeutic Sponge        and Method of Making, issued Mar. 29, 1949.    -   ⁴Correll, et al., U.S. Pat. No. 2,507,244, Surgical Gelatin        Dusting Powder and Process for Preparing Same, issued May 9,        1950.    -   ⁵Studer, et al., U.S. Pat. No. 2,558,395, Undenatured Gelatin        Hemostatic Sponge Containing Thrombin, issued Jun. 26, 1951.    -   ⁶Sieger, et al., U.S. Pat. No. 2,899,362, Hemostatic Sponges and        Method of Preparing Same, issued Aug. 11, 1959.    -   ⁷Song, et al., U.S. Pat. No. 5,399,361, Collagen-containing        Sponges as Drug Delivery Compositions for Proteins, issued Mar.        21 1995.

⁸Cragg, et al., U.S. Pat. No. 6,071,301, Device and Method forFacilitating Hemostasis of a Biopsy Tract, issued Jun. 6, 2000.

-   -   ⁹Cragg, et al., U.S. Pat. No. 6,086,607, Device and Method for        Facilitating Hemostasis of a Biopsy Tract, issued Jul. 11, 2000.    -   ¹⁰Cragg, et al., U.S. Pat. No. 6,162,192, System and Method for        Facilitating Hemostasis of Blood Vessel Punctures with        Absorbable Sponge, issued Dec. 19, 2000.    -   ¹¹Pawelchak, et al., U.S. Pat. No. 4,292,972, Lyophilized        Hydrocolloid Foam, issued Oct. 6, 1981.    -   ¹²Sawyer, U.S. Pat. No. 4,238,480, Method for Preparing an        Improved Hemostatic Agent and Method of Employing the Same,        issued Dec. 9, 1980.    -   ¹³Sawyer, U.S. Pat. No. 4,404,970, Hemostatic Article and Method        for Preparing and Employing the Same, issued Sep. 20, 1983.

All of the above references are herein incorporated by reference intheir entirety to the same extent as if each individual reference wasspecifically and individually indicated to be incorporated by referencein its entirety.

3. State of the Art

Cross-linked gelatin, often in the form of gelatin foam, gelatin film orgelatin sponges, has been used as a hemostatic agent since itsdevelopment by Correll in 1945.sup.1-4 Such hemostatic sponges or foamsare used routinely to pack wounds, absorb blood and stop bleeding. Theyare often left in place to be bioabsorbed over a period of weeks tomonths. In addition, medicaments, such as antibiotics, growth factorsand thrombus enhancing agents, have been incorporated into thecross-linked gelatin to enhance the in vivo properties of thecomposition.sup.5-7

To use, the hemostatic sponge is placed onto or into the wound,whereupon it absorbs blood or other fluids, expands to pack and compressthe wound, and initiates a rapid clotting response. However, to achievecomplete fluid absorption, rapid expansion, effective wound compression,and vigorous clotting action, the sponge must first be pre-hydrated orwetted, usually with sterile saline, before placement in the wound. Thispre-hydration step, often accompanied with gentle compression and/ormassaging of the sponge, can be time consuming and troublesome, oftenyielding non-uniform results at a critical moment in patient care. (See,for example, Directions for Use provided with Gelfoam®—available fromPharmacia Upjohn).

The hydrated sponge can then be compressed by hand and inserted ortamped into the wound. One particular wound suitable for use by hydratedsponges is a puncture site such as a puncture wound resulting fromcatheter insertion or a biopsy needle. When so used, the art describesejection of a pledget of cross-linked gelatin from a syringe into thepuncture site.sup.8-10

Critical to the ejection process is the flowability of the pledget fromthe syringe assembly and retention of its structural integrity duringinsertion into the body. Specifically, ejection of the pledget from thesyringe assembly is preferably conducted with, at most, moderatepressure to ensure accurate placement in vivo. Efficient ejection of thepledget, in turn, relates to the flowability of the cross-linked gelatinthrough the syringe assembly. Higher fluid content pledgets are believedto correlate with enhanced flowability and, accordingly, it is desirableto maintain as high a fluid absorbability content in the sterilizedpledget as possible.

Likewise, it is critical that the structural integrity of the pledget issubstantially maintained as it is ejected from the syringe assembly whenplaced in vivo in order to ensure that portions of the pledget are nottorn or otherwise separated from the pledget. This criticality isparticularly important when the pledget is placed over a blood vesselpuncture in order to avoid unintended thrombosis of the vessel.Structural integrity of the gelatin composition of the pledget underpressure is believed to correlate with the tensile strength of thecomposition and, accordingly, it is desirable to maintain as high atensile strength in the sterilized pledget as possible.

In addition, it is desirable to perform hydration of the pledgets in thesterile environment of an injection syringe or syringe assembly byaddition of a sterile saline solution to the syringe. Such a hydrationprocess is complicated by the potential of distortions in the pledgetdue to a lack of uniformity and consistency in the rate of hydration ofthe pledget. Such distortions could cause unanticipated difficulties infitting the pledget into the identified wound, e.g., puncture. Inaddition, the length of time normally needed to hydrate the pledgetwould typically prohibit such a hydration process. Accordingly, it isdesired to obtain a quicker, more uniform hydration of the pledget toensure predictability of the size and shape of the pledget and tofacilitate hydration of the pledget inside the injection syringe.

SUMMARY OF THE INVENTION

This invention is directed to a biocompatible, hemostatic, cross-linkedgelatin composition which has a wetting agent incorporated therein. Inparticular, this invention is directed to a biocompatible materialcomprising a cross-linked gelatin and a sufficient amount of wettingagent to permit uniform wetting of the gelatin in the presence of anaqueous solution. This invention further relates to methods of usingthis composition to decrease the hydration time of a hemostatic,cross-linked gelatin composition. This invention further relates to akit of parts for preparing a biocompatible, hemostatic, cross-linkedgelatin composition comprising a sterile syringe and non-hydrated,biocompatible, pledget comprising a cross-linked gelatin and a wettingagent.

Applicants have surprisingly and unexpectedly found that wetting agentscan be incorporated into a cross-linked gelatin composition, improvingtheir use as, e.g., a hemostatic sponge.

Applicants have further surprisingly found that the incorporation ofwetting agents into a hemostatic, cross-linked gelatin compositiondecreases the hydration time needed to prepare the gelatin for use.Incorporation of the wetting agent into the cross-linked gelatincomposition provides for one or more of improved fluid absorption,gelatin expansion and wound compression.

Applicants have also surprisingly found that hemostatic, cross-linkedgelatin compositions with wetting agents incorporated therein are moreuniformly lubricated when ejected as a pledget from a syringe assemblyand less likely to suffer damage to the structural integrity of thepledget. Such pledgets are also more quickly hydrated, allowing forhydration of the pledget inside the injection syringe. Further, thepledgets made according to the invention are more uniformly hydrated,and less likely to become distorted during the hydration process.

Accordingly, in one embodiment, this invention is directed to abiocompatible, hemostatic, cross-linked gelatin composition comprising across-linked gelatin and a sufficient amount of wetting agent to permituniform wetting of the gelatin in the presence of an aqueous solution.

In one embodiment, the incorporation of the wetting agent into thecross-linked gelatin can be achieved by mixing or impregnating thegelatin with wetting agent prior to foaming. In this embodiment, thewetting agent can comprise from about 0.01 to 10 weight percent of thegelatin and preferably from about 0.1 to 10 weight percent.

In another embodiment, the incorporation of the wetting agent into thecross-linked gelatin can be achieved by coating the wetting agent overthe surface of a gelatin sponge. In this embodiment, a solution ofwetting agent and liquid solvent carrier is prepared wherein the wettingagent comprises from about 0.01 to 20 percent of the solution andpreferably from about 1 to 10 weight percent. The solution is coatedover the surface of the sponge, then the liquid solvent carrier isevaporated. After evaporation, the wetting agent comprises from about0.01 to about 5 weight percent of the gelatin sponge and preferably fromabout 0.1 to about 5 weight percent.

In both embodiments, the biocompatible, hemostatic sponge can bebioabsorbable. The sponge can also include one or more compositionsselected from the group consisting of growth factors, thrombus enhancingagents, and antimicrobial agents.

In one of its method aspects, the invention is directed to a method fordecreasing the hydration time of a cross-linked gelatin compositionwhich method comprises, prior to hydration of said cross-linked gelatin,incorporating a biocompatible wetting agent with said cross-linkedgelatin.

The biocompatible, hemostatic sponge of the present invention can besterilized and packaged to facilitate use in surgical procedures.

Accordingly, in another of its aspects, this invention is directed to akit of parts for preparing a biocompatible, hemostatic, cross-linkedgelatin composition comprising a syringe or syringe assembly and anon-hydrated pledget, said pledget comprising a cross-linked gelatin andwetting agent.

BREIF DESCRIPTION OF THE DRAWINGS

FIG 1. is a side view of a hemostatic compound delivery system.

DETAILED DESCRIPTION OF THE INVENTION

This invention is directed to a biocompatible, hemostatic, cross-linkedgelatin composition which has a wetting agent incorporated therein. Inparticular, this invention is directed to a biocompatible materialcomprising a cross-linked gelatin and a sufficient amount of wettingagent incorporated therein to permit uniform wetting of the sponge inthe presence of an aqueous solution and to methods of using thiscomposition to decrease the hydration time of the gelatin composition.Prior to describing this invention in further detail, the followingterms will first be defined.

Definitions

As used herein, the following terms have the following meanings:

The term “cross-linked gelatin” refers to well known gelatin foams orsponges which are cross-linked with a conventional cross-linking agentsuch as formaldehyde as described in the art by Correll.sup.1-3 The term“cross-linked gelatin composition” refers to compositions comprisingcross-linked gelatin. Such compositions often include other componentssuch as a medicament.sup.8-10 or a second polymer such ascollagen.sup.13 and starch.sup.6.

The term “growth factors” refers to those medicaments which areconventionally employed to facilitate tissue growth such as theendothelial wall of a punctured blood vessel or biopsy tract. Examplesof suitable growth factors include PDGF, EGF, FGF, IGF's 1 and 2, TGFand the like.

The term “thrombus enhancing agents” refers to those medicaments whichare conventionally employed to facilitate thrombus formation at apuncture site such as at the endothelial wall of a punctured bloodvessel or a biopsy tract. Examples of suitable thrombus enhancing agentsinclude thrombin, fibrinogen, Factor XIII and other coagulation factors.

The term “antimicrobial agent” refers to agents which destroy microbes(i.e., bacteria, fungi, viruses and microbial spores) thereby preventingtheir development and pathogenic action. Preferred antimicrobial agentsinclude antibiotics and antiviral agents and, in particular,antibiotics. Other preferred antimicrobials include, for example, iodinecontaining materials such as PVP-I.sub.2, chlorhexidene, and the like.

The term “packaging element” refers to those packaging components usedto encase the cross-linked gelatin and include, by way of example,boxes, syringes, envelopes, tubings, catheters, introducers and thelike. The packaging elements may comprise glass, plastic, ceramics,cardboard, paper products and the like.

The term “wetting agent” refers to a biocompatible agent whichfacilitates or enhances the hydration or lubrication of a hemostaticsponge. Examples of suitable wetting agents include polyoxyalkylenes(such as BASF Pluronics, UCC Carbowaxes, PEGs), ether cappedpolyoxyalkylenes, e.g., polyoxyethylene lauryl ether, ester cappedpolyoxyalkylenes, e.g., polyoxyethylene stearate, sorbitan esters (suchas certain products called Span and Tween), phosphatides (such aslecithin), alkyl amines, glycerin, water soluble polymers such aspolyethylene oxides, carboxymethyl cellulose, polyvinyl alcohol, andpolyvinyl pyrrolidone, surfactants such as alkyl (C.sub.6-C.sub.20)sulfate salts, e.g., sodium lauryl sulfate, aryl (C.sub.6-C.sub.10)sulfate salts, and alkaryl (C.sub.7-C.sub.24) sulfate salts, and thelike.

Compositions

The compositions of this invention include biocompatible hemostatic,cross-linked gelatin compositions comprising a cross-linked gelatincomposition and a sufficient amount of wetting agent to permit uniformwetting of the gelatin in the presence of an aqueous solution.

The wetting agent is utilized in an amount sufficient to enhance thehydration and/or lubrication of the composition. When the wetting agentis incorporated into the cross-linked gelatin, the wetting agent willcomprise from about 0.01 to about 10 weight percent based on the totalweight of the composition and preferably from about 0.1 to 10 weightpercent. When the wetting agent is coated onto the surface of thecross-linked gelatin, the wetting agent will comprise from about 0.01 toabout 20 weight percent, based on the total weight of the appliedsolution (preferably from about 1 to 20 weight percent), and 0.01 to 5weight percent based on the total weight of the gelatin compositionafter evaporation of the liquid solvent of the applied solution (andpreferably from about 0.1 to 5 weight percent).

In a preferred embodiment, the cross-linked gelatin composition furthercomprises a medicament such as an antimicrobial agent (e.g., anantibiotic), growth factors, thrombus enhancing agents, and the like ora property modifying agent such as a wetting agent. Mixtures ofmedicaments and property modifying agents can also be used. Suitablemedicaments can be mixed with or impregnated into the cross-linkedgelatin composition prior to sterilization procedures.

When employed, the medicament is utilized in an amount sufficient forits intended purpose, e.g., an antimicrobially effective amount, anamount sufficient to induce thrombus formation, and an amount sufficientto promote growth. The specific amount employed relates to theeffectiveness of the medicament, the disease condition of the patientbeing treated, the age and weight of the patient, the location of thedisease and other factors well within the purview of the attendingclinician.

Prior to administration of the biocompatible, hemostatic, cross-linkedgelatin to the patient, the gelatin will be sterilized. Sterilizationcan be achieved by, for example, heat or E-beam sterilization.Biocompatible wetting agents will typically be incorporated into orcoated onto the cross-linked gelatin composition prior to thesterilization procedure. E-beam sterilization of a cross-linked gelatincomposition is described in U.S. Provisional Patent Application Ser. No.60/275,391, filed Mar. 12, 2001, entitled “Methods for SterilizingCross-Linked Gelatin Compositions” (Attorney Docket No. 032005-093),which application is incorporated herein by reference in its entirety.Heat sterilization of a cross-linked gelatin composition is described inU.S. Pat. No. 2,465,357.sup.3

In the embodiment, described in FIG. 1, a pledget of the cross-linkedgelatin composition of the present invention can be inserted into asyringe assembly 10 comprising a holding chamber 14, an injection portwhich comprises a luer hub 20, and an ejection port 22 which is attachedto a cannula 24. The syringe assembly containing the pledget 16 can bepackaged and sterilized. Then, sterile saline 12 can be added to theholding chamber 14 in a sufficient amount to hydrate the pledget 16.Once the pledget is hydrated, the pledget 16 can be transferred into thecannula 24 attached to the ejection port 22.

The incorporation of wetting agents into the cross-linked gelatincomposition can allow for rapid, uniform wetting of the gelatin andelimination of the pre-hydration process. The wetting agent can be addedand dissolved (or dispersed) directly into the hemostat chemicalformulation, preferably just before the foaming process. An amount ofwetting agent ranging from about 0.01 to 10 percent based on the weightof the gelatin material, and preferably from 0.1 to 10 percent, can beused. Processes for manufacturing a hemostatic sponge are disclosed inU.S. Pat. No. 2,465,357.sup.3 and U.S. Pat. No. 2,507,244.sup.4.

Alternatively, the wetting agent could be dissolved (or dispersed) in anon-aqueous liquid solvent (e.g., ethanol, isopropanol, acetone and thelike) coated onto the cross-linked gelatin composition and the liquidsolvent removed by evaporation. In another embodiment, the wetting agentand liquid solvent mixture could be applied onto the gelatin compositionand the foam compressed. The liquid solvent would then be removed byevaporation. When the wetting agent is coated over the gelatincomposition, the concentration of the wetting agent in the liquidsolvent is preferably from about 0.01 to 20 percent and preferably from1 to 20 weight percent and more preferably 1 to 10 weight percent.

Suitable biocompatible wetting agents are commercially available andinclude, for example, sodium lauryl sulfate, Pluronic F-68, PluronicF-38, Pluronic P-105, Pluronic-10R5, Tween 20, Tween 60, Tween 85, Brij35, Brij 78, Myrj 52, PEG 600, glycerin and the like.

In addition, biocompatible wetting agents can be incorporated intobiocompatible collagen hemostatic sponges (e.g., Actifoam™) to improvetheir wetting times in the manner and concentrations set forth above.Suitable wetting agents include Tween 20 (1% in isopropanol) andPluronic P-105(1% in isopropanol).

EXAMPLES

The following examples are set forth to illustrate the claimed inventionand are not to be construed as a limitation thereof.

In these examples, unless otherwise indicated, all temperatures are indegrees Centigrade and all percents are weight percents based on thetotal weight of the composition. Similarly, the following abbreviationsare employed herein and are as defined below. Unless defined, theabbreviations employed have their generally accepted meaning:

cc=cubic centimeter

cm=centimeter

EtOH=ethanol

g=gram

IPA=isopropyl alcohol

min.=minute

mm=millimeter

mL=milliliters

pcs=pieces

RH=relative humidity

RO water=reversed osmosis water

RPM=rotations per minute

sec=seconds

wt=weight

Example 1 Comparison of Hydration Time of Cross-Linked Gelatin withWetting Agent Incorporated Therein Vs. Cross-Linked Gelatin withoutWetting Agent

The purpose of this example is to demonstrate the feasibility ofdifferent agents in reducing the hydration time of a cross-linkedgelatin having a variety of candidate wetting agents incorporated intothe gelatin prior to foaming (Gelatin Composition 1A). In this example,the surfactant was added to the solution used to prepare thecross-linked gelatin foam.

The results of the amount of time required to wet the gelatin wascompared to the hydration time of a cross-linked gelatin which does nothave any candidate wetting agents incorporated therein (ComparativeGelatin Composition 1B).

A. Preparation of Gelatin Composition 1A

Gelatin Composition 1A was prepared as follows. A stock solution of 5%pigskin gelatin in purified bottled water was prepared by heating to60.degree. C. with constant stirring. The solution was cooled to37.degree. C. and 0.01% formalin added. This was incubated for twohours. To produce a 10% surfactant to gelatin ratio (i.e., 10% loading),1.5 grams of surfactant was dissolved in 20 grams of water, and thissolution added to 280 gram aliquots of the stock gelatin solution.

Foam was produced by whipping air into the warm mixture for about 2minutes using a kitchen blender. The whipping ceased when the foampeaked. The gelatin foam was then transferred to screens and containedwithin metal shells until firm, about 10-15 minutes. The foam was driedat 32.2.degree. C. (90.degree. F.) and 18% RH, which took about 24-36hours. Test samples were prepared by cutting the skin from the driedfoam buns and cutting cubes of about 1.5-2.0 cm from central locationsin the bun. For additional information regarding the preparation of suchgelatin compositions, see U.S. Pat. Nos. 2,465,357 and 2,507,244.

The following candidate wetting agents were each incorporated into thegelatin prior to foaming:

-   1. Polyacrylamide (weight average molecular weight of about 1500)    (as a sample of an cationic wetting agent)-   2. Sodium lauryl sulfate (as a sample of a anionic wetting agent)-   3. Pluronic F-68 (poly(ethylene oxide)-co-(propylene oxide) block),    (as a sample of a non-ionic wetting agent—number average molecular    weight of about 8400) (commercially available from BASF Corp.)-   4. Tween 20 (polyoxyethylene sorbitan monolaurate), (as a sample of    a non-ionic wetting agent—molecular weight of approximately 1227)-   5. PEG (600) (poly(ethylene glycol), (as a sample of a non-ionic    wetting agent—molecular weight of about 600)-   6. Glycerin (as a sample of a non-ionic wetting agent)

B. Preparation of Comparative Gelatin Composition 1B

Comparative Gelatin Composition 1B was prepared as above except foromission of the wetting agent.

C. Comparison of the Foam Properties and Hydration Time of Cross-LinkedGelatin with Wetting Agent Incorporated Therein Vs. Gelatin WithoutWetting Agent

(1) Foam Properties

Most of the gelatin compositions produced a smooth, white, meringue-likefoam, which dried into a high, rigid, compressible bun.

Gelatin Compositions 1A produced with lauryl sulfate, Pluronic F-68,Tween 20 and 30% lauryl sulfate candidate wetting agents were softer andless compressible. These compositions also displayed some loss of foamheight and an increase in cell size and voids. The Tween 20 and 30%lauryl sulfate formulations demonstrated the most collapse and increasein cell size and voids.

(2) Hydration

Table 1 sets forth the hydration times for the Gelatin Compositions 1Aand the Comparative Gelatin Compositions. Hydration was tested in bothuncompressed and compressed (comp.) foams. The concentration ofcandidate wetting agent in each composition is indicated as a percentloading. Specifically, the percent loading refers to the weight percentof the wetting agent based on gelatin in the composition before foamingbased on a 300 mL solution containing 5% gelatin.

The Gelatin Compositions 1A with the lauryl sulfate (30% loading), Tween20 and Pluronic F-68 wetting agents yielded a startling reduction inhydration time of about 3 to 10 seconds (versus 6 minutes for theComparative Gelatin Composition 1B (control)). The amount of waterabsorbed was somewhat reduced for the 30% lauryl sulfate containingsample. It was basically unchanged for the other two compositions.

Unless otherwise noted, all samples were the aggregate of 3 pieces.

TABLE 1 Hydration Times for Gelatin Compositions 1A and ComparativeGelatin Compositions Dry Wet Hydration Weight Weight Wet/Dry WettingAgent Time (min) (g) (g) (3 pcs) Comparative Gelatin 6.5-7.0 0.13 3.7429 Composition 1B (0% loading) Comparative Gelatin 6.0-6.5 0.12 4.23 35Composition 1B (0% loading) (Comp 15 - 2 mm) Gelatin Composition 1A >200.14 ND ND Polyacrylamide Wetting Agent (10% loading) GelatinComposition 1A >20 0.16 ND ND Polyacrylamide Wetting Agent (10% loading)(Comp 15 - 2 mm) Gelatin Composition 1A 45 sec 0.24 4.30 18 Laurylsulfate Wetting Agent (10% loading) Gelatin Composition 1A 45 sec 0.447.55 17 Lauryl sulfate Wetting Agent (10% loading) (Comp 20 - 2 mm)Gelatin Composition 1A 10-15 sec 0.18 3.01 17 (1 pc) Lauryl sulfateWetting Agent (30% loading) Gelatin Composition 1A 10 sec 0.17 3.04 18(1 pc) Lauryl sulfate Wetting Agent (30% loading) (Comp 15 - 1 mm)Gelatin Composition 1A 3-5 sec 0.17 4.13 24 Pluronic F-68 Wetting Agent(10% loading) Gelatin Composition 1A 3.5 sec 0.15 3.73 25 Pluronic F-68Wetting Agent (10% loading) (Comp 15 - 3 mm) Gelatin Composition 1A 5sec 0.37 10.90  29 Tween 20 Wetting Agent (10% loading) GelatinComposition 1A 5 sec 0.28 8.08 29 Tween 20 Wetting Agent (10% loading)(Comp 10 - 2 mm) Gelatin Composition 1A 4.5 0.18 5.15 29 PEG 600 WettingAgent (10% loading) Gelatin Composition 1A 3.5 0.22 6.01 27 PEG 600Wetting Agent (10% loading) (Comp 20 - 2 mm) Gelatin Composition 1A 4.50.25 5.13 21 Glycerin Wetting Agent (10% loading) Gelatin Composition 1A3.5 0.25 6.71 27 Glycerin Wetting Agent (10% loading) (Comp 20 - 2 mm)Comparative Gelatin 5.5 0.16 4.72 30 Composition 1B (0% loading)

The wet/dry ratio reported is the wet weight over the dry weight whichprovides a ratio of the amount of water absorbed per unit weight of drymaterial.

The above data indicates that all of the candidates used, with theexception of polyacrylamide, act as wetting agents per this invention.On the other hand, the increase in wetting time provided by usingpolyacrylamide indicates that this material is not a suitable wettingagent.

The above data of the different agents used, little or no change in thewet/dry ratio occurred with the use of either Tween 20 or PEG 600;modest lowering of the wet/dry ratio occurred with the use of PluronicF-68 and Glycerin; and somewhat greater lowering of the wet/dry ratiooccurred with the use of lauryl sulfate.

Example 2 Comparison of Hydration Times Cross-Linked GelatinCompositions with Wetting Agent Coating Vs. Cross-Linked GelatinCompositions without a Wetting Agent Coating

The purpose of this example is to demonstrate the reduction in hydrationtime of a cross-linked gelatin composition containing differentnon-ionic wetting agents (Gelatin Composition 2A) and forming asurfactant coating on the preformed cross-linked gelatin foamcomposition. Specifically, in this example, the wetting agents werecoated onto the cross-linked gelatin composition in the followingmanner: the wetting agent was added to a solvent, the cross-linkedgelatin composition was added to the resulting solution, the soak timeused was 10 seconds or 1 minute, as indicated below, the cross-linkedgelatin composition was removed, excess liquid drained, and then thesolvent was allowed to evaporate.

The hydration time required to wet these compositions was compared tothe hydration time of a cross-linked gelatin composition without such awetting agent coating (Comparative Gelatin Composition 2B (no coating)and Comparative Gelatin Composition 2C (carrier solvent coating)).

A. Preparation of Gelatin Composition 2A

Gelatin Composition 2A was prepared as follows. First, ComparativeGelatin Composition 1B, as set forth in Example 1, was prepared.

Second, ten percent solutions of each of the following wetting agentswere prepared in isopropanol (2-Propanol, ACS reagent (Aldrich Cat.#19076-4)) in 30 cc screw cap vials.

-   -   1. Tween 20 (polyoxyethylene (20) sorbitan monolaurate) (Aldrich        Cat. #27434-8)    -   2. Tween 60 (polyoxyethylene (20) sorbitan monostearate)        (Aldrich Cat. #37425-3)    -   3. Tween 85 (polyoxyethylene (20) sorbitan trioleate) (Aldrich        Cat. #38890-4)    -   4. Brij 35 (polyoxyethylene (23) lauryl ether) (Aldrich Cat.        #85836-6) (Sigma)    -   5. Myrj 52 (polyoxyethylene (40) stearate) (Aldrich Cat. #P3440)        (BASE)    -   6. Brij 78 (polyoxyethylene (23) steryl ether) (Sigma #23600-4)

A 2×0.2×0.0.6 cm piece of Gelatin Composition 2A was placed into eachvial and the vial inverted to soak the composition with the solution.This was repeated for each solution. Two contact/soak times wereemployed, 10 seconds and one minute.

The coated Gelatin Compositions 2A were then removed, drained of excessliquid and air dried overnight.

B. Preparation of Comparative Gelatin Composition 2B and 2C

Comparative Gelatin Composition 2B (untreated) was prepared as set forthfor Comparative Gelatin Composition 1B in Example 1.

Comparative Gelatin Composition 2C (carrier solvent coating) wasprepared as follows. A 2.times.2.times.0.6 cm piece of ComparativeGelatin Composition 1B (as set forth in Example 1) was placed into a 30cc screw cap vial of isopropanol and the vial inverted to soak thecomposition with the solution. Two contact/soak times were employed, 10seconds and one minute. After soaking, the gelatin compositions werethen air dried overnight.

C. Comparison of Hydration Times

Dry samples of each of the Gelatin Compositions 2A and the ComparativeGelatin Compositions 2B and 2C were compressed to about 0.1 cm anddropped into a 250 mL beaker containing 100 mL of RO water.

The time for each gelatin composition to reach full hydration wasmeasured with a digital stopwatch. Hydrated samples were weighed andreweighed to assure full hydration. Hydration was also determined bymonitoring when the composition turned from opaque to translucent.

The results of this evaluation are shown in Table 2. Where two valuesare given, duplicate runs were made.

TABLE 2 Hydration Times for Gelatin Composition 2A and ComparativeGelatin Compositions 2B and 2C Soak Time in Wetting Agent HydrationWet/Dry Gelatin Composition Solution Time Wt. Comparative Gelatin 0 6, 6min 28, 33 Composition 2B Gelatin Composition 2A 10 sec 20, 25 sec 49,35 Tween 20 Wetting Agent Gelatin Composition 2A 1.0 min 15, 20 sec 49,58 Tween 20 Wetting Agent Gelatin Composition 2A 10 sec 3, 1.5 min 41,44 Tween 60 Wetting Agent Gelatin Composition 2A 1.0 min 24, 35 sec 38,31 Tween 60 Wetting Agent Gelatin Composition 2A 10 sec 30, 30 sec 41,45 Tween 85 Wetting Agent Gelatin Composition 2A 1.0 min 35, 35 sec 46,34 Tween 85 Wetting Agent Gelatin Composition 2A 10 sec 15, 20 sec 38,26 Myrj 52 Wetting Agent Gelatin Composition 2A 1.0 min 40, 40 sec 37,29 Myrj 52 Wetting Agent Gelatin Composition 2A 10 sec 15, 15 sec 55, 42Brij 35 Wetting Agent Gelatin Composition 2A 1.0 min 40, 40 sec 52, 33Brij 35 Wetting Agent Gelatin Composition 2A 10 sec 15, 15 sec 38, 39Brij 78 Wetting Agent Gelatin Composition 2A 1.0 min 20, 20 sec 43, 42Brij 78 Wetting Agent Comparative Gelatin 10 sec 3 min 40 Composition 2CComparative Gelatin 1.0 min 3 min 40 Composition 2C

The wet/dry ratio reported is the wet weight over the dry weight whichprovides a ratio of the amount of water absorbed per unit weight of drymaterial.

All samples of cross-linked gelatin, treated or not, yielded very largeand similar uptakes for water (wet/dry weight). Also, as evident above,the best wetting agents were deemed those that produced the most rapidhydration, e.g., Tween 20, Brij 35 and Brij 78.

Example 3 Demonstration of Improved Hydration Time for Cross-LinkedGelatins with Wetting Agent Coatings Containing Ethanol and IsopropanolCarrier Solvents

The purpose of this example is to demonstrate that the improvedhydration time achieved for cross-linked gelatins coated withnon-aqueous wetting agents as per Example 2 above were independent ofthe solvent used to coat the gelatin.

Specifically, gelatin composition 3A was prepared in the same manner asgelatin composition 2A. The wetting agent coating solution comprises aselected wetting agent and a carrier solvent of either ethanol orisopropanol. Soak contact time of the foam in the wetting solution wasabout 10 seconds each.

After the coating layer has dried, the gelatin compositions werecompressed and soaked in a 250 mL beaker containing 100 mL of RO water.The hydration times were measured as in Example 2.

The specific wetting agents employed were as follows:

-   -   1. Tween 20 (polyoxyethylene (20) sorbitan monolaurate) (Aldrich        Cat. #27434-8)    -   2. Pluronic F-68 (poly(ethylene oxide)-co-(propylene oxide)        block), (as a sample of a non-ionic wetting agent—average        molecular weight of about 8400) (commercially available from        BASF Corp.)    -   3. Pluronic P-105 (poly(ethylene oxide)-co-(propylene oxide)        polymer), (as a sample of a non-ionic wetting agent—average        molecular weight of about 6500) (commercially available from        BASF Corp.)    -   4. Pluronic-10R5 (poly(ethylene oxide)-co-(propylene oxide)        polymer), (as a sample of a non-ionic wetting agent—average        molecular weight of about 4550) (commercially available from        BASE Corp.).    -   5. Pluronic-F38 (poly(ethylene oxide)-co-(propylene oxide)        block), (as a sample of a non-ionic wetting agent—average        molecular weight of about 5000) (commercially available from        BASE Corp.).

Hydration times for the compositions are set forth in Table 3 below.Where multiple values are given, multiple runs were made.

TABLE 3 Hydration Times for Gelatin Compositions 3A Wetting Agent Usedin Gelatin Carrier Composition 3A Solvent Hydration Times Tween 20 3%ETOH 10, 15; 10, 15, 15 sec Tween 20 3% IPA 20, 20; 10, 10, 15 sec Tween20 10% ETOH 10, 10 sec Tween 20 10% IPA 10, 10 sec F-38 3% ETOH 40, 40,40 sec F-38 3% IPA 30, 45, 45 sec F-68 3% ETOH 30, 45, 45 sec F-68 3%IPA 20, 25, 20 sec P-105 3% ETOH 10, 15, 10 sec P-105 3% IPA 10, 10, 10sec 10R5 3% ETOH 10, 20, 20 sec 10R5 3% IPA 10, 10, 10 sec

The average wet/dry ratio for the above samples was approximately 44with a range of from 39-56 wherein this ratio is calculated by the wetweight over the dry weight which provides a ratio of the amount of waterabsorbed per unit weight of dry material.

The above data demonstrates that there is no appreciable difference inthe hydration time of the coated cross-linked gelatin using differentsolvents to coat the gelatin.

However, the solvent employed should be non-aqueous to the extent thatwater irreversible damages the foam. Preferably, the non-aqueous solventis ethanol and/or isopropanol since methanol causes some degradation ofthe foam properties and acetone exhibits a disagreeable odor.

Example 4 Hydration Times for Cross-Linked Gelatin Compositions withWetting Agent Coating of Varying Concentrations

This example examines the effect of wetting agent concentration onhydration time used to coat the cross-linked gelatin composition.

Specifically, cross-linked gelatin compositions with wetting agentcoatings were prepared as in Example 2. The wetting agent coatingsolutions were prepared with concentrations of the selected wettingagents of 0.1, 1.0, 3.0 and 10.0%. The hydration times were measured oncompressed samples, in the manner set forth in Example 2.

All of the wetting agents and concentrations demonstrated rapidhydration times, although at the highest concentration of 10%, thehydration time was somewhat reduced. The results are set forth in Table4 below. Where multiple values are given, multiple runs were made.

TABLE 4 Hydration Times for Cross-Linked Gelatin Compositions withWetting Agent Coatings of Varying Concentrations Wetting Agent UsedHydration Times at Wetting Agent Concentration in Coating 0.10% 1.00%3.00% 10.00% Tween 20 15, 10; 60, 15, 10; 15, 20, 10 sec 20, 10 sec 25,15 sec 10, 10; 10 sec Brij 35 10, 10, 10; 15, 10, 10; 10, 20, 15 20, 30,30 35, 20, 20 sec 30, 20 sec sec sec Brij 78 10, 15, 10 sec 10, 10; 25sec 10, 10 sec P-105 15, 15 sec 10, 10; 10 sec 10, 10 sec 10R5 15, 15sec 10, 10 sec 10, 10 sec

The wet/dry ratio reported is the wet weight over the dry weight whichprovides a ratio of the amount of water absorbed per unit weight of drymaterial.

The above data demonstrates that the concentration of wetting agentemployed in the coating solution does not materially affect thehydration time of the coated cross-linked gelatin.

Example 5 Hydration Times for Cross-Linked Gelatin Compositions withWetting Agent of Varying Concentrations Incorporated Therein

This example evaluates the effect of wetting agent concentration usedduring the formation of a gelatin foam composition on both the hydrationtime and the wet/dry ratio. This example differs from the previousexample in that the wetting agent is added to the composition prior tofoam formation as per Example 1 rather than coating a prior formedcross-linked gelatin foam with the wetting agent as per Examples 2-4.

Specifically, cross-linked gelatin compositions with wetting agentsincorporated therein were prepared as in Example 1 except that 0.02weight percent of formalin was used and wherein the wetting agent wasadded to the gelatin composition prior to foam formation. The wettingagent coating solutions were prepared with concentrations set forth inTable 5.

The hydration times for these gelatin compositions were measured bymanually compressing each cut sample and dropping it into a beakercontaining 150 mL of tap water. Hydration was complete when the foamturned translucent and were measured on compressed samples, in themanner set forth in Example 2. Hydration time of a control, similar toComparative Gelatin Example 2B, was also measured.

The hydration times are set forth in the Table 5 below.

TABLE 5 Wetting Concen- Foam Hydration Wt. Agent tration Wet Foam DryTime Wet/Dry Tween 20 5% ¾ in. ⅜ in. 2-3 sec NA Tween 20 3% ½ in. ⅜ inc.2-3 sec 23.4 Tween 20 1% ¾ in. ¼ in. 2-3 sec 24.0 Tween 20 0.10%   ¾ in.⅝ in. 5 sec 41.5 Tween 60 3% no foam NA NA Tween 85 3% no foam NA NAMyrj 52 3% no foam NA NA Brij 35 3% 1¼ in. ¼ in. 2-3 sec 16.5 Brij 78 3%½ in. ⅜ in. 2-3 sec 12.0 F-68 3% 1 in. ¼ in. 2-3 sec 19.7 Control 0% 1¾in. 1¼ in. 8 min 45.8

These results indicate that gelatin foam compositions with wettingagents did not achieve the same level of foam formation as compared tothe control. Upon drying, foam heights for these compositions werefurther reduced, with the lowest concentration of surfactant causingless height reduction.

The compositions with the wetting agents incorporated therein alsotended to be softer and more resilient, more fibrous and with largerirregular cells than the control compositions. Again, the lowestconcentration of surfactant (0.1%) gave foams closer to the control.

All gelatin foams with wetting agent incorporated therein demonstratedvery rapid and complete hydration. Hydration was complete within 2-3seconds for the surfactant gelatin samples compared to about 8 minutesfor the control. At the 0.1% level, the presence of the surfactant wassufficient to dramatically improve hydration times.

Unfortunately, water uptake was significantly diminished for allsurfactant samples, except those with the lowest concentration ofsurfactant. Mechanical strength of all the samples (wet or dry) wasexcellent, and equal to that of the control.

Based on the above data, when the wetting agent is incorporated prior tocross-linked gelatin foam formation, very low concentrations of wettingagent are preferred. Moreover, the data indicates minimal impact of lowconcentrations of wetting agent on reduced hydration time indicatingthat such low concentrations advantageously provide for a compositioncloser to control in many properties but surprisingly have excellenthydration times as compared to control.

Preferred wetting agents as demonstrated in this example utilizing theaddition of wetting agent prior to foam formation should not preventfoam sponge formation of the cross-linked gelatin.

Example 6

This example demonstrates that the incorporation of a wetting agent intoa gelatin composition does not deleteriously effect the clotting abilityof the gelatin composition as compared to control. A partialthromboplastin test (PTT) was employed. The PTT assay is a generalscreening assay for the detection of coagulation abnormalities in theintrinsic coagulation pathway. This is particularly important because,notwithstanding improved hydration time, a cross-linked gelatincomposition having an adverse effect on blood clotting would becontra-indicated for use, e.g., as a hemostatic sponge.

Specifically, in this example, fresh whole human blood was drawn atAppTec Laboratory Services using vacutainer tubes containing sodiumcitrate. The citrated whole blood was spun down. The plasma wasseparated from the red blood cells, pooled into a polypropylene testtube, and stored on ice. This plasma was used as the exposure medium fortesting.

The comparison sample, test tube, reference material and controls wereplaced in polypropylene tubes and exposed to plasma (determined by the 1mL/4 cm.sup.2 exposure ratio) for 15 minutes at 37.+-0.1.degree. C. inan agitating water bath at 60 RPM. The volume of 0.3 mL of plasma wasused to saturate the test and comparison articles prior to exposure. Atthe end of the incubation period, the plasma was removed from thematerials. The plasma samples were placed on ice and tested on theCascade M4.

The results are set forth in Tables 6 and 7 below.

TABLE 6 Exposure Ratios Coagulation Time (seconds) Surface Assay % ofNeg Article Area (cm²) (mL) Values Ave. Control Cross-linked 4.0 1.0300/226.7/300 275.6 92 Gelatin - Ex. 2B Gelfoam 4.0 1.0 300/300/300 300100 (Pharmacia- Upjohn) Positive 4.0 1.0 NA 39.9 13 Control, BlackRubber Reference 4.0 1.0 NA 174.3 58 Material, Latex Tubing Negative NA1.0 NA 300 NA Control, Human Plasma

TABLE 7 Exposure Ratios Coagulation Time (seconds) Surface Assay % ofNeg Article Area (cm²) (mL) Values Ave. Control Cross-linked 4.0 1.0300/208.2/300 269.4 90 Gelatin - Ex. 2A (0.3% Tween 20) Gelfoam 4.0 1.0300/300/300 300 100 (Pharmacia- Upjohn) Positive 4.0 1.0 NA 39.9 13Control, Black Rubber Reference 4.0 1.0 NA 174.3 58 Material, LatexTubing Negative NA 1.0 NA 300 NA Control, Human Plasma

If coagulation does not occur within 300 seconds, the result is reportedas “300 sec.”).

The above data demonstrates that the compositions of this inventionexhibit substantially equivalent effects on clotting time as compared tocontrol and, accordingly, it was concluded that these compositions arecompatible for use as, e.g., hemostatic sponges.

Example 7

This example evaluates the histopathologic results of a total of 2arterial femoral puncture sites from one pig, using the compositions ofthis invention. The objective of this evaluation was to assess the acuteand chronic outcome of femoral puncture sites following closure with thecompositions of this invention delivered into the tissue tract adjacentto the vessel puncture using a cannula delivery tube. The puncture woundin the right femoral artery was sealed with cross-linked gelatincomposition comprising 0.1% Tween 20 obtained from solvent coating ofthe composition per Example 2 above. The puncture wound in the leftfemoral artery was sealed with cross-linked gelatin compositioncomprising 0.3% Tween 20 obtained from solvent coating of thecomposition per Example 2 above. Subsequently, approximately 2-4 weekslater, the animal was sacrificed, the arteries adjacent the femoralpuncture site were excised and fixed in formalin.

Specifically, the formalin-fixed arterial explants were trimmed in thearea of interest every 3 mm into 11 to 14 segments (sections 1 to 14).The sections' sequence was started at the proximal end of each explant.The sections were embedded in paraffin, sectioned at approximately 5microns three times serially, and stained with Hematoxylin and Eosin(HE), Masson's Trichrome (MT), and Verhoeffs Van Gieson (VVG) stain,respectively. Histologic evaluation was performed and data wererecorded.

A 1 to 5 grading scale was utilized, where 1 is minimal, 2 is mild, 3 ismoderate, 4 is marked, and 5 is severe. Loss of endothelium was gradedaccording to circumferential spread of the change. The change ofendothelium loss spanned approximately up to 20% of the circumference ingrade 1, 20-40% in grade 2, 40-60% in grade 3, 60-80% in grade 4 andmore than 80% in grade 5. Necrosis of the media was graded according todepth extension. The change of media necrosis extended intoapproximately up to 20% of the media thickness in grade 1, 20-40% ingrade 2, 40-60% in grade 3, 60-80% in grade 4 and more than 80% in grade5.

1-Animal 3094G—Right Femoral Artery

Gross Observations: The segment of artery was 52 mm long. There were nosignificant gross changes in the intact explant. Trimming revealedsub-occlusive thrombosis in most sections (1 through 8) and partialthrombosis in sections 9 and 10. The delivery tract could not be seengrossly.

Microscopic Observations: The puncture site was observed in sections 9and 10 (24 and 27 mm from origin). There was transmural laceration withregeneration and fibrosis of the edges. From these lacerated sites andthe corresponding intimal areas of more proximal sections a partiallyorganized thrombus projected into the lumen and was nearly occluding thelumen. There was mineralization of the media in a sub-intimal position(sections 1, 2, 7, 8, 9), due to localized media pressure necrosis(dystrophic mineralization). Mineralization was also observed in theorganizing thrombus. There was slight to mild partial loss ofendothelium in the proximal sections (sections 1-7) that was consideredto be the result of erosion associated with termination catheterization.In approximately the proximal ⅔ of the explant (sections 1-10) there wasfibrosis of the media, most likely due to pressure necrosis at the timeof treatment. The implant was not observed. There was mature connectivetissue in the adventitia possibly corresponding to the treatment tract.These microscopic observations are consistent with arterial needlepuncture and healing.

2-Animal 3094G-Left Femoral Artery

Gross Observations: The segment of artery was 50 mm long. There were nosignificant gross changes in the intact explant.

Microscopic Observations: Transmural laceration (puncture site) wasobserved in section 8 (18-21 mm, recut 3) and was characterized bybifocal laceration across the vessel with proliferation of fibrousgranulation tissue across the lumen dividing the pre-existing arterialwall into two symmetrically placed remnants. The residual lumen wasconfined to one of the arterial remnants. The lumen was constructed(moderate stenosis) by intimal fibrous proliferation at the puncturesite. There was mineralization of the external elastic lamina adjacentto the laceration site. There was moderate fibrosis in the adventitia atthe laceration site and in a separate area in the more distantadventitial tissue that most likely corresponds to the puncture tract.There was no residual implant material and there was no significantinflammatory response in the tract. There was minimal to mild loss ofendothelium that was considered to be the result of erosion associatedwith termination catheterization.

The access tract produced by the catheter was found in the vascular wallin both samples as a localized fibrosis in the adventitia and focaltransmural fibrosis. The left artery displayed bifocal transmurallaceration and healing by fibrous connective tissue and mediaregeneration bridging across the lumen, indicating focal puncturethrough opposite sides of the vessel. There was no significant occlusivethrombosis in the left artery but focal moderate stenosis associatedwith intimal hyperplasia at the puncture site was present. The rightartery there was sub-occlusive thrombosis that proceeded from thepuncture site and extended proximally. There was evidence of pronouncedarchitectural disruption and media necrosis due to mechanical trauma tothe vessel wall at the puncture site, believed to be responsible forthrombosis. No residual foam test material was observed. Thesemicroscopic observations are consistent with arterial needle punctureand healing.

The above data demonstrates that the compositions of this invention arewell tolerated in vivo.

Example 8

This example evaluates the cytotoxicity of the compositions of thisinvention as compared to prior art compositions, i.e., in the absence ofa wetting agent. In this example, samples were tested in the ISO agaroseoverlay cytotoxicity test—a test well known in the art for determiningcytotoxicity potential. Specifically, samples were placed onto petridishes, in triplicate, containing agarose the surface of which containsa monolayer of L-929 mouse fibroblast cells which are sensitive tocytotoxic agents. Subsequently, the samples were evaluated forreactivity and grade depending upon the amount of cellular damage ordeath directly beneath and/or surrounding the material. The scoring isbased on the following:

0=no zone under or around sample

1=some malformed or degenerated cells under sample

2=zone limited to area under sample

3=zone extends 0.5 to 1.0 cm beyond sample

4=zone extends greater than 1.0 cm

In this example, a score of 0-2 is deemed to be non-toxic; a score of 3is deemed to be weakly or moderately toxic; and a score of 4 is stronglytoxic.

The results of this evaluation are summarized below in Table 8:

TABLE 8 Sample Score Cross-linked gelatin w/o surfactant 0 Cross-linkedgelatin with Tween 20 (prepared by Example 2 - isopropanol) 0.3% Tween20 0 1.0% Tween 20 1, 1 (two triplicate runs) Cross-linked gelatin withPluronic P-105 (prepared by Example 2 - isopropanol) 0.3% Pluronic P-1050, 0 (two triplicate runs) 1.0% Pluronic P-105 0, 0 (two triplicateruns)

The above results demonstrate that the compositions of this invention,as tested in this example, are non-cytotoxic.

From the foregoing description, various modifications and changes in theabove described methods will occur to those skilled in the art. All suchmodifications coming within the scope of the appended claims areintended to be included therein.

The invention claimed is:
 1. A kit of parts for preparing abiocompatible, hemostatic, cross-linked gelatin composition comprising:a sterile syringe assembly; a saline solution; a non-hydrated pledget,said pledget comprising a pre-formed cross-linked gelatin sponge coatedby soaking the non-hydrated pledget in a non-aqueous solvent having awetting agent dissolved therein; wherein evaporation of the non-aqueoussolvent coats at least a substantial portion of the surface of thenon-hydrated pledget with the wetting agent; wherein the wetting agentdecreases hydration time of the non-hydrated cross-linked gelatinpledget.
 2. The kit of claim 1, wherein the syringe assembly comprises aholding chamber, an ejection port, and a cannula.
 3. The kit of claim 2,wherein the syringe assembly is adapted to combine the saline solutionand the non-hydrated pre-formed cross-linked gelatin pledget includingthe wetting agent coated on at least a substantial portion of thesurface thereof in the holding chamber and subsequently eject thecross-linked gelatin pledget.
 4. The kit of claim 1, wherein the coatingsolution comprises from about 0.01 to about 20 weight percent of thewetting agent.
 5. The kit of claim 1, wherein the coated non-hydratedpre-formed cross-linked gelatin pledget comprises from about 0.01 toabout 5 weight percent of the wetting agent.
 6. The kit of claim 5,wherein the coated non-hydrated pre-formed cross-linked gelatin pledgetis bioabsorbable.